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A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample
BACKGROUND: Bread wheat has a large complex genome that makes whole genome resequencing costly. Therefore, genome complexity reduction techniques such as sequence capture make re-sequencing cost effective. With a high-quality draft wheat genome now available it is possible to design capture probe se...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5899405/ https://www.ncbi.nlm.nih.gov/pubmed/29653520 http://dx.doi.org/10.1186/s12864-018-4640-y |
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author | Olohan, Lisa Gardiner, Laura-Jayne Lucaci, Anita Steuernagel, Burkhard Wulff, Brande Kenny, John Hall, Neil Hall, Anthony |
author_facet | Olohan, Lisa Gardiner, Laura-Jayne Lucaci, Anita Steuernagel, Burkhard Wulff, Brande Kenny, John Hall, Neil Hall, Anthony |
author_sort | Olohan, Lisa |
collection | PubMed |
description | BACKGROUND: Bread wheat has a large complex genome that makes whole genome resequencing costly. Therefore, genome complexity reduction techniques such as sequence capture make re-sequencing cost effective. With a high-quality draft wheat genome now available it is possible to design capture probe sets and to use them to accurately genotype and anchor SNPs to the genome. Furthermore, in addition to genetic variation, epigenetic variation provides a source of natural variation contributing to changes in gene expression and phenotype that can be profiled at the base pair level using sequence capture coupled with bisulphite treatment. Here, we present a new 12 Mbp wheat capture probe set, that allows both the profiling of genotype and methylation from the same DNA sample. Furthermore, we present a method, based on Agilent SureSelect Methyl-Seq, that will use a single capture assay as a starting point to allow both DNA sequencing and methyl-seq. RESULTS: Our method uses a single capture assay that is sequentially split and used for both DNA sequencing and methyl-seq. The resultant genotype and epi-type data is highly comparable in terms of coverage and SNP/methylation site identification to that generated from separate captures for DNA sequencing and methyl-seq. Furthermore, by defining SNP frequencies in a diverse landrace from the Watkins collection we highlight the importance of having genotype data to prevent false positive methylation calls. Finally, we present the design of a new 12 Mbp wheat capture and demonstrate its successful application to re-sequence wheat. CONCLUSIONS: We present a cost-effective method for performing both DNA sequencing and methyl-seq from a single capture reaction thus reducing reagent costs, sample preparation time and DNA requirements for these complementary analyses. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12864-018-4640-y) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-5899405 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-58994052018-04-23 A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample Olohan, Lisa Gardiner, Laura-Jayne Lucaci, Anita Steuernagel, Burkhard Wulff, Brande Kenny, John Hall, Neil Hall, Anthony BMC Genomics Methodology Article BACKGROUND: Bread wheat has a large complex genome that makes whole genome resequencing costly. Therefore, genome complexity reduction techniques such as sequence capture make re-sequencing cost effective. With a high-quality draft wheat genome now available it is possible to design capture probe sets and to use them to accurately genotype and anchor SNPs to the genome. Furthermore, in addition to genetic variation, epigenetic variation provides a source of natural variation contributing to changes in gene expression and phenotype that can be profiled at the base pair level using sequence capture coupled with bisulphite treatment. Here, we present a new 12 Mbp wheat capture probe set, that allows both the profiling of genotype and methylation from the same DNA sample. Furthermore, we present a method, based on Agilent SureSelect Methyl-Seq, that will use a single capture assay as a starting point to allow both DNA sequencing and methyl-seq. RESULTS: Our method uses a single capture assay that is sequentially split and used for both DNA sequencing and methyl-seq. The resultant genotype and epi-type data is highly comparable in terms of coverage and SNP/methylation site identification to that generated from separate captures for DNA sequencing and methyl-seq. Furthermore, by defining SNP frequencies in a diverse landrace from the Watkins collection we highlight the importance of having genotype data to prevent false positive methylation calls. Finally, we present the design of a new 12 Mbp wheat capture and demonstrate its successful application to re-sequence wheat. CONCLUSIONS: We present a cost-effective method for performing both DNA sequencing and methyl-seq from a single capture reaction thus reducing reagent costs, sample preparation time and DNA requirements for these complementary analyses. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12864-018-4640-y) contains supplementary material, which is available to authorized users. BioMed Central 2018-04-13 /pmc/articles/PMC5899405/ /pubmed/29653520 http://dx.doi.org/10.1186/s12864-018-4640-y Text en © The Author(s). 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Methodology Article Olohan, Lisa Gardiner, Laura-Jayne Lucaci, Anita Steuernagel, Burkhard Wulff, Brande Kenny, John Hall, Neil Hall, Anthony A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample |
title | A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample |
title_full | A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample |
title_fullStr | A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample |
title_full_unstemmed | A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample |
title_short | A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample |
title_sort | modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic dna sample |
topic | Methodology Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5899405/ https://www.ncbi.nlm.nih.gov/pubmed/29653520 http://dx.doi.org/10.1186/s12864-018-4640-y |
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